1. Diagnostic criteria
The pathological diagnosis of lung cancer in biopsy tissue specimens is mainly to clarify the presence or absence of tumor and tumor type. For patients with advanced inoperable disease, the pathological diagnosis should be subtype classification as far as possible, and for cases with atypical morphology, immunohistochemical staining should be combined. The diagnosis of “non-specific type” should be avoided as much as possible. Biopsies from patients with advanced NSCLC should also be examined for molecular pathology, especially in patients with adenocarcinoma. The histologic type of lung cancer in large surgically resected specimens should be based on the most recent version of the WHO lung cancer classification. Pathologic diagnosis of adenocarcinoma in situ, microinvasive adenocarcinoma and large cell carcinoma cannot be made in small biopsy specimens, intraoperative frozen specimens, and requires surgical resection of the entire specimen or adequate sampling of the tumor to make a definitive diagnosis.
2. Diagnostic guidelines
The pathological diagnosis guideline of lung cancer consists of specimen processing, specimen sampling, pathological examination and pathological report, etc.
(1) Key points of specimen processing: 10 neutral buffered formaldehyde fixative is recommended, avoid using fixative containing heavy metals, the volume of fixative should be ≥10 times the volume of the fixed specimen, and fixation at room temperature. Specimens should not be fixed for more than 60 minutes from isolation. Biopsy specimens should be placed directly into the fixative, and lobar or whole lung resection specimens can be fixed by injecting a sufficient amount of fixative from the bronchus, or by inserting a probe along the bronchial wall and tumor incision lung tissue. Fixation time: 6 to 24 hours for small biopsy specimens; 12 to 48 hours for surgical resection specimens.
Cytology smear (sputum, pleural effusion) fixation should be used 95 ethanol fixative, the time should not be less than 15 minutes, or non-gynecologic liquid-based cytology fixative (fixation time and method can be operated according to the instructions); when the need to make exfoliated cell wax block, the cell mass after centrifugation and tissue fixation procedures are the same, using 10 neutral buffered formaldehyde fixative, the time ≥ 2 hours.
(2) General description of the specimen and requirements for sampling
(1) Biopsy specimens were checked for accuracy and all tissues sent for examination were taken.
(2) Local pneumonectomy specimens
(1) Remove surgical sutures or metal staples.
②Record the size of the specimen and the pleural surface.
The size of the mass, its surface condition (with or without hemorrhage, necrosis, or cavity formation), its relationship to the pleura and lung parenchyma, and the distance between the edge of the mass and the cut edge are described.
The tumor, tumor and pleura, and tumor and lung parenchyma margins should be cut according to the location and size of the lesion, and the entire tumor should be taken when the tumor is <3 cm.
3) Lobectomy specimens
①Examine the five basic structures of the lung: airways, lung parenchyma, pleura, blood vessels, and lymph nodes. The size of the specimen should be measured and the lung hilum should be used to position the specimen.
②Take the bronchial margins, vascular margins and the closest part of the tumor to the pleura, or the adhesions to other lung lobes.
③Find the hilar lymph nodes.
④According to the location and status of the tumor, there are 2 options: first, to cut the specimen along the bronchial wall and the tumor through the lung tissue (which can be done with the help of a probe inserted into the trachea) and open the bronchi and their branches in order to best expose the structural relationship between the lesion and the bronchi at all levels and the surrounding lung tissue. Second, for specimens injected with formaldehyde in the main bronchus, incisions should be made at 0.5 to 1.0 cm intervals, and the sections should be coronal and perpendicular to the hilum.
⑤ Describe the size of the tumor, the cut surface (with or without hemorrhage, necrosis, or cavity formation), its location within the lobes and segments of the lung and its relationship to the bronchi, the extent of the lesion (focal or metastatic), and distal or local secondary changes. The number of blocks to be obtained depends on the size of the specific lesion (all tumors <3 cm should be obtained), the specific location, the presence of concomitant lesions (related to clinical staging), and should include the tumor and the pleura, the tumor and the lobe or segmental bronchus (varies by specimen), the tumor and the surrounding lung or secondary lesions, the tumor and the lung section or bronchial section, etc. The cross-lobe specimen should also include the part of the tumor in relation to the cross-lobe. All lymph nodes in N2 or other areas should be counted for clinical examination. The recommended volume of the tissue block should not be larger than 2.5×1.5
The recommended volume of the tissue block is not larger than 2.5×1.5×0.3cm.
(3) Key points of pathological description: The general description includes specimen type, tumor size, relationship with bronchus (different types of specimens) or pleura, other concomitant lesions or multiple lesions, and cut edges.
The diagnosis includes tumor site, histologic subtype, extent of involvement (bronchus, pleura, vasculature, nerves, types of concomitant lesions, intrapulmonary foci, lymph node metastases, etc.), margins, and special staining, immunohistochemistry, or molecular pathology results as necessary. The information included should meet the needs of clinical staging and give pTNM staging. For multiple lung cancers, the nature of the lesion should be clarified as much as possible based on the morphological characteristics of each lesion, i.e., intrapulmonary metastatic cancer or multiple primary cancers.
(4) Immunohistochemistry, special staining and molecular pathology: TTF-1, Napsin-A, p63, p40 and CK5/6 should be used as immunohistochemical markers to differentiate adenocarcinoma from squamous carcinoma, and only TTF-1 and p40 can be selected if there is insufficient tissue; CD56, Syn, CgA, Ki-67 and TTF-1 should be used as neuroendocrine tumor markers. The diagnosis of neuroendocrine tumor can be made only if at least one neuroendocrine marker is clearly positive and the number of positive cells is more than 10 tumor cells based on the morphological characteristics of neuroendocrine tumor.
For patients with stage II-IIIA NSCLC, N1/N2 positive non-squamous carcinoma and small specimen squamous carcinoma, mutation of epidermal growth factor receptor (EGFR) in tumor tissue is recommended. For patients with advanced NSCLC, EGFR mutations, anaplastic lymphoma kinase (ALK), ROS1 and RET fusion genes, and CMET exon 14 jump mutations should be routinely performed along with the diagnosis of tumor tissue. For those who are eligible, tests for mutations in KRAS, BRAF, HER2, NTRK1/2/3 and NRG1/2 fusion genes are available. For immunotherapy, PD-L1 immunohistochemistry can be performed; EGFR mutations can be detected by amplification blocked mutation system or high-throughput sequencing (HTSHTS); ALK fusion genes can be detected by Ventana immunohistochemistry, FISH, RT-PCR or HTS. Detection of ROS1 fusion genes can be performed by RT-PCR, FISH or HTS; RET gene fusions and CMET exon 14 jump mutations are preferred and recommended to be detected together with other driver gene detection, either by RT-PCR or HTS. In patients with advanced NSCLC where tissue is not available, blood can be used as a complement to tissue for EGFR testing, and highly sensitive amplified blocked mutation systems, HTS, or digital PCR can be used; for ALK, ROS1, RET fusion genes, and CMET exon 14 jump mutations, liquid biopsy specimens are not recommended as a first step. EGFRT790M testing is recommended for patients with resistance to EGFR TKIs. Histology is the gold standard, and blood ctDNA EGFR T790M testing can be a useful supplement when tissue is not available.
3. Diagnostic pathology report
(1) Tumor
①Tissue typing (including morphological subtypes)
②The extent of involvement
③Whether it invades the pleura
④Vascular infiltration
⑤Nerve invasion
(2) Cutting edge
①Bronchial margin
②Vascular margin
(3) Lung margins (local lung margin specimens)
(3) Other pathological findings (e.g., obstructive pneumonia, treatment-related changes, etc.)
(4) Regional lymph nodes (including peribronchial, hilar and separate lymph nodes)
①Total number
②Number of involved lymph nodes
(5) Distant metastases
(6) Other tissues/organs
(7) pTNM staging
(8) Difficult cases submitted to higher hospitals for consultation (provide the original pathology report to verify the information of the submitted sections to reduce errors, provide adequate lesion sections or wax blocks, and intraoperative observations, etc.)